Skeletal dysplasia-causing mutations in TRPV4 alter the chondrocyte’s transcriptomic response to mechanical loading

Transient receptor potential vanilloid 4 (TRPV4) is a mechanosensitive ion channel highly expressed in chondrocytes that maintains cartilage homeostasis during development and throughout aging. Mutations in the channel cause a wide range of developmental disorders including skeletal dysplasias. The gain-of-function mutations V620I and T89I have been identified to lead to brachyolmia and metatropic dysplasia, respectively. Although it is known these mutations suppress hypertrophic differentiation, the mechanisms by which they alter chondrocyte response to mechanical loading remain to be elucidated. To determine the effect of these mutations on chondrocyte mechanotransduction, tissue-engineered cartilage was derived from differentiated CRISPR-edited human induced pluripotent stem cells harboring the moderate V620I or severe T89I TRPV4 mutations. Tissue-engineered hiPSC-derived cartilage was then subjected compressive mechanical loading at physiological levels, and RNA-sequencing was used to assess the transcriptomic signatures of wildtype and mutant tissue-engineered cartilage. Our results demonstrate that the V620I and T89I mutations lead to reduced mechanoresponsiveness, characterized by a lower number of differentially expressed genes and diminished activation of genes downstream of TRPV4 signaling and contributing towards the process of endochondral ossification. Changes in extracellular matrix production and organization were found to contribute towards the phenotype in V620I chondrocytes, whereas dysregulated retinoic acid signaling was linked to T89I, and disrupted proliferation was common to both. Our findings suggest that dysfunctional mechanotransduction arising due to the V620I and T89I mutations contribute to the developmental phenotypes observed in their respective skeletal dysplasias, introducing potential pharmacologic targets for these conditions.